Session Information

to investigate the expression of Thymic Stromal Lymphopoietin (TSLP) in diffuse cutaneous systemic sclerosis (dcSSc) patients and explore its effects in vivo and in vitro comparing with IL-13 and TGF-beta stimulations.

Methods:

Skin biopsies [dcSSc; n=13 and healthy controls (HC); n=12] were used for immunohistochemistry (IHC) and immunofluorescence studies using TSLP, CD4+, CD8+, CD31+, and CD163+ markers. Wild type (WT) and IL4Ra1-deficient mice (IL4Ra1-ko) were treated with TGF-beta, IL-13, Poly(I:C), or TSLP. Human fibroblasts and peripheral blood mononuclear cells (PBMCs) were stimulated with the same cytokines. Gene expression (microarray and rt-PCR) and protein levels of phospho-Smad2 were tested.

Results:

TSLP was highly expressed in skin of dcSSc patients, stronger in perivascular areas, where we observed inflammatory cell infiltrates, and in interstitial cells. TSLP expression was co-localized with immune cells, such as CD4+, CD8+, although mainly produced by CD163+ cells. Skin of TSLP-treated mice showed upregulated clusters of genes that overlapped with IL-13 and TGFb-treated mice. In addition, a specific TSLP-cluster showed upregulation of CXCL9, proteasomes, and other interferon-regulated genes. In PBMCs, TSLP alone upregulated mannose-receptor-1 (MRC1), an alternatively-activated macrophage marker, to a similar degree as after IL-13 stimulation. MRC1 was also highly expressed in dcSSc skin compared to controls. TSLP kinetics, along 24 hours of stimulation in PBMCs, showed an early induction of TNF, MX1, and INFg, followed by an induction of CXCL9 and MRC1 gene-expression. Human fibroblasts and skin of mice-treated with TSLP showed TGF-beta-canonical pathway activation with phosphorylation of Smad2. The lack of IL4Ra1 in TSLP-treated mice promotes similar cutaneous inflammation and upregulation of profibrotic markers (PAI-1 and CXCL5). Poly(I:C)-treated mice, a SSc-like murine model, showed high levels of TSLP in similar areas as seen in the skin of dcSSc patients and also mainly in infiltrating immune cells, shown by IHC.

Conclusion:

TSLP is highly expressed in skin of dcSSc patients, mainly produced by macrophages, and regulates similar genes as other profibrotic cytokines (TGF-beta and IL-13), strongly suggesting that it promotes SSc fibrosis directly or by stimulating production or activation of these cytokines. TSLP also promotes a proinflammmatory effect, which might explain this dual finding in SSc patients.